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Lab report 7.docx - Microbiology Lab HSCI2003-2019 Reports...

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Microbiology LabHSCI2003-2019ReportsTitle3.7-3.9 Isolation of Actinomyces from soil and theeffect of antibiotics on bacterial growthNameRochelle YuLab Partners: Ivory ChenGroup7Date Performed2019/10/29 and 2019/11/5
1. AbstractThe main objective of the first experiment was isolation, purification, and to understandmechanism of antibiotics production of actinomyces. Soil samples were serially diluted andplated on actinomycete isolation agar media.In the second experiment, heat shocktransformation and antibiotics selection are carried out in order to understand the commonmechanism of which antibiotics inhibit bacterial growth. As for the purpose of the Kirby-Bauerdisk diffusion susceptibility test is to determine the sensitivity or resistance of pathogenic aerobicand facultative anaerobic bacteria to various antimicrobial compounds. The pathogenic organismis grown on Mueller-Hinton agar in the presence of various antimicrobial impregnated filterpaper disks. The presence or absence of growth around the disks is an indirect measure of theability of that compound to inhibit that organism.2. Material and MethodsExperiment 3.7Materials:-Dry soil from campus-Gauze’s synthetic medium-0.1% potassium dichromate / phenol / Eppendorf-Sterilized water and saline-Culture broth (Escherichia coli, Alcaligenes faecalis, Bacillus megaterium)-Sterilized discs-Spreader and forcepsMethods:1.Soil from the campus are collected as samples.2.The samples are then packed in a sterilized kraft paper bag and stored for 2 weeks atroom temperature.3.After two weeks, 5g of soil samples were transferred into a sterilized flask and washeated at 120oC for 15mins.
4.For the treated group, phenol and sterilized water are added into the flask, the solutionsare mixed on the shaker for 30mins, then was left to stand for 5mins. As for the controlgroup, water and soil are added into the flask (without heat and phenol).5.Three agar plates are labelled as the treated group (1, 10-1), while the remain three platesare labelled as the control group(1, 10-1).6.Dilutions were prepared by adding sample into the saline solution. Original or dilutedsamples were then spread onto the agar plates.7.Lastly, the plates were incubated for 1 to 2 weeks. The color, number of colonies, and thedifference between control and treated were examined for further analysis.Experiment 3.8Materials:-DNA plasmid-Sterile water-Chemical competent cells on ice-LB broth-LB agar plates with ampicillin-SpreaderMethods:1.Two agar plates are first labelled with group number and plasmid number.2.The DNA plasmid and sterile water are added onto the plates aseptically.3.The mixture of DNA and competent cells on ice are incubated for 30mins.4.The cells were then heat shocked at 42oC for 20secs with no shaking and was transferredback to the ice for 2mins. The cells are recovered at 37o

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